title: Activation of chimeric and full-length growth hormone receptors by growth hormone receptor monoclonal antibodies. A specific conformational change may be required for full-length receptor signaling.
abstract: Signal transduction by the growth hormone receptor (GHR) occurs through growth hormone (GH)-induced dimerization of two GHRs to form a trimeric complex. It is thought that dimerization alone is sufficient for signaling, since monoclonal antibodies (mAbs) against the extracellular domain of the GHR elicit proliferation of FDC-P1 cells transfected with a chimeric receptor comprising the extracellular domain of the GHR and the fibronectin and cytoplasmic domains of the murine granulocyte colony-stimulating factor receptor. We have screened 14 GHR mAbs for proliferative activity against characterized FDC-P1 and BaF-B03 cell lines stably expressing the full-length human, rabbit, or rat GHR, or the chimeric human GHR/granulocyte colony-stimulating factor receptor, and for transactivation of the c-fos promoter and STAT activation. With the chimeric receptor, eight mAbs were able to elicit proliferation, although there was no correlation between inhibition of hormone binding and agonist activity. In contrast, no mAbs were able to act as agonists with the full-length GHR FDC-P1 cell lines, although nine competed with GH for binding. A weak proliferative response was observed in the BaF-B03 cell lines with two of the mAbs (263 and 1C9), and the addition of anti-mouse F(ab)2 resulted in increased signaling in the hGHR BaF-B03 cell line to a plateau of 28 +/- 4% of the GH maximum for mAb 263. These data could indicate considerable stringency in the ability of mAbs to correctly dimerize the full-length GHR. However, the ability of mAb 263 to stimulate a mutant hGHR altered in the F'-G' loop of domain 2 was nearly abolished, concurrent with an increased affinity of this mAb for the receptor. Since the F'-G' loop undergoes a conformational change on GH binding and is necessary for full proliferative signaling, we propose that in addition to promoting receptor dimerization, mAb 263 may induce specific changes in receptor conformation similar to GH, which are required for the biological response.
authors.affiliation: Department of Physiology and Pharmacology and Centre for Molecular and Cellular Biology, University of Queensland, St. Lucia, Queensland 4072, Australia.
authors.forename: S W
authors.initials: SW
authors.lastname: Rowlinson
authors.forename: S N
authors.initials: SN
authors.lastname: Behncken
authors.forename: J E
authors.initials: JE
authors.lastname: Rowland
authors.forename: R W
authors.initials: RW
authors.lastname: Clarkson
authors.forename: C J
authors.initials: CJ
authors.lastname: Strasburger
authors.forename: Z
authors.initials: Z
authors.lastname: Wu
authors.forename: W
authors.initials: W
authors.lastname: Baumbach
authors.forename: M J
authors.initials: MJ
authors.lastname: Waters
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal / Q000494:pharmacology; D002455:Cell Division; D002460:Cell Line; D004305:Dose-Response Relationship, Drug; D006801:Humans; D010446:Peptide Fragments / Q000819:agonists / Q000737:chemistry / Q000235:genetics / Q000276:immunology; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D011487:Protein Conformation; D016760:Proto-Oncogene Proteins c-fos / Q000235:genetics; D011817:Rabbits; D051381:Rats; D016188:Receptors, Granulocyte Colony-Stimulating Factor / Q000819:agonists; D011986:Receptors, Somatotropin / Q000819:agonists* / Q000737:chemistry / Q000235:genetics / Q000276:immunology*; D011993:Recombinant Fusion Proteins / Q000819:agonists / Q000737:chemistry / Q000276:immunology; D015398:Signal Transduction; D015533:Transcriptional Activation
chemical_list: D000911:Antibodies, Monoclonal; D010446:Peptide Fragments; D016760:Proto-Oncogene Proteins c-fos; D016188:Receptors, Granulocyte Colony-Stimulating Factor; D011986:Receptors, Somatotropin; D011993:Recombinant Fusion Proteins